Flow Sorting of Marine Bacterioplankton after Fluorescence In Situ Hybridization

Sekar, Raju; Fuchs, Bernhard M.; Amann, Rudolf; Pernthaler, Jakob

doi:10.1128/aem.70.10.6210-6219.2004

Cited by 139 publications

(108 citation statements)

References 61 publications

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“…15 to 29% of total counts in the Baltic Sea ), while they are often absent from pelagic marine systems, including sites in the Southern Ocean ) and offshore Arctic Ocean ). However, recent molecular studies have identified marine β-Proteobacteria, notably phylotypes of Nitrosomonas and Nitrosospira (Nold et al 2000, Freitag & Prosser 2004, Sekar et al 2004) at polar ocean sites . β-Proteobacteria have been recorded in lake snow (Weiss et al 1998) and in the freshwater melt pools on Arctic pack ice (Brinkmeyer et al 2004).…”

Section: Discussionmentioning

confidence: 99%

Prokaryotic community structure and heterotrophic production in a river-influenced coastal arctic ecosystem

Garneau

Vincent²,

Alonso‐Sáez³

et al. 2006

Aquat. Microb. Ecol.

131

105

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Spatial patterns in prokaryotic biodiversity and production were assessed in the Mackenzie shelf region of the Beaufort Sea during open-water conditions. The sampling transect extended 350 km northwards, from upstream freshwater sites in the Mackenzie River to coastal and offshore sites, towards the edge of the perennial arctic ice pack. The analyses revealed strong gradients in community structure and prokaryotic cell concentrations, both of which correlated with salinity. Picocyanobacterial abundance was low (10 2 to 10 3 cells ml -1 ), particularly at the offshore stations that were least influenced by the river plume. Analysis by catalyzed reporter deposition for fluorescence in situ hybridization (CARD-FISH) showed that the dominant heterotrophic cell types were β-Proteobacteria at river sites, shifting to dominance by α-Proteobacteria offshore. Cells in the Cytophaga-Flavobacter-Bacteroides and γ-Proteobacteria groups each contributed < 5% of total counts in the river, but >10% of counts in the marine samples. Archaea were detected among the surface-water microbiota, contributing on average 1.3% of the total DAPI counts in marine samples, but 6.0% in turbid coastal and riverine waters. 3 H-leucine uptake rates were significantly higher at 2 stations influenced by the river (1.5 pmol l -1 h -1 ) than at other marine stations or in the river itself (≤0.5 pmol -1 h -1 ). Size-fractionation experiments at 2 coastal sites showed that > 65% of heterotrophic production was associated with particles > 3 µm. These results indicate the importance of particleattached prokaryotes, and imply a broad functional diversity of heterotrophic microbes that likely facilitates breakdown of the heterogeneous dissolved and particulate terrestrial materials discharged into arctic seas. KEY WORDS:Prokaryote diversity · Archaea · Proteobacteria · Cytophaga-FlavobacterBacteroides · Arctic Ocean · Mackenzie River estuary · Picocyanobacteria · CARD-FISH Resale or republication not permitted without written consent of the publisherAquat Microb Ecol 42: [27][28][29][30][31][32][33][34][35][36][37][38][39][40] 2006 ments (Payette et al. 2004) may mobilize the large stocks of organic carbon contained within their soils and cause increased transfer of these materials to arctic rivers and ultimately to the ocean. Heterotrophic microbiota are likely to play a major role in the response of coastal arctic ecosystems to ongoing change, but prokaryotic diversity and production in these cold ocean environments have been little explored (Amon 2004).In the western Canadian Arctic, the Mackenzie River discharges large quantities of freshwater, solutes and sediments into a vast shelf region of the Beaufort Sea that extends >100 km offshore and encompasses a total area of 63 600 km 2 (Carmack et al. 2004). The annual discharge of the Mackenzie River (330 km 3 yr -1 ; Macdonald et al. 1998) is the 4th highest in the Arctic Basin after the Siberian rivers Yenisei, Lena and Ob, with concomitantly large inputs of freshwater biota, terrestria...

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Section: Discussionmentioning

confidence: 99%

Prokaryotic community structure and heterotrophic production in a river-influenced coastal arctic ecosystem

Garneau

Vincent²,

Alonso‐Sáez³

et al. 2006

Aquat. Microb. Ecol.

131

105

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“…on undisturbed samples of aquifer material, using the fluorescence in situ hybridisation (FISH) method Swiger and Tucker 1996;Wagner et al 2003). It is also possible to use FISH to label specific microbial cells and then sort and count the cells by means of flow cytometry (Sekar et al 2004).…”

Section: Methodsmentioning

confidence: 99%

Review: Microbial biocenoses in pristine aquifers and an assessment of investigative methods

2006

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The current knowledge of microbial biocenoses (communities) in pristine aquifers is presented in a review, which also discusses their relevance for questions of groundwater protection. Aquifers are heterogeneous on all scales and structured in a variety of habitats. The void spaces in many aquifers are small. The biocenoses are thus predominantly composed of microorganisms and, often, microinvertebrates. Larger voids and macroorganisms occur in karst cavities. Due to the absence of light, the biocenoses depend on chemical energy resources, which are, however, scarce in non-contaminated groundwater. The microorganisms thus show small cell sizes, low population densities and reduced activity; they developed specific strategies to survive oligotrophic conditions. The review also discusses the impact of contamination on the biocenoses, and the potential use of the biocenoses or specific organisms as indicators for groundwater quality, and the limits of this approach. Bacteria are either planktonic or attached to aquifer material, which requires both fluid and solid phase sampling. Most groundwater bacteria are viable but non-culturable. Consequently, cultivation techniques give an incomplete picture of the biocenoses, while methods from molecular microbiology provide genetic fingerprints of the entire community. Different analytical methods are available to count microorganisms, identify species, characterise microbial diversity, and measure activity.Résumé Cette revue expose l'état actuel des connaissances concernant les biocénoses microbiennes présentes dans les aquifères oligotrophes. L'impact d'une contamination sur les biocénoses est discuté, ainsi que le potentiel que Les bactéries sont planctoniques ou attachées aux matériaux de l'aquifère, ce qui demande unéchantillonnagè a la fois de l'eau et du substrat. De nombreuses méthodes sont aujourd'hui disponibles pour le comptage, l'identification et la caractérisation de la diversité, ainsi que la mesure des activités des organismes des aquifères. Comme la grande majorité des bactéries est viable mais non cultivable, les techniques de cultures actuelles ne donnent qu'une image incomplète des communautés, alors que les méthodes moléculaires développées récemment offrent la possibilité d'obtenir un profil de la communauté plus complet.Resumen Se presentauna reseña crítica del conocimiento actual de biocenosis microbiana (comunidades) en acuíferos prístinos la cual también discute su relevancia en términos de protección de aguas subterráneas. Los acuíferos son heterogéneos en todas las escalas y estructurados en una variedad de habitats. Los espacios vacíos en muchos acuíferos son pequeños. La biocenosis está por lo tanto compuesta predominantemente por microorganismos y, frecuentemente, microinvertebrados. Espacios más grandes y macroorganismos ocurren en cavidades kársticas. Debido a la ausencia de luz la biocenosis depende de recursos energéticos químicos los cuales, sin embargo, son escasos en agua subterránea no contaminada. Los microorganismos muestran...

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“…An example of this for environmental analysis is combined FISH and Fluorescence Activated Sorting (FISH-FACS) for the detection and sortingbased enrichment of specific cell types from natural samples. Kalyuzhnaya et al (62) used this approach to examine methanotrophic bacteria from lake sediments and Sekar et al (63) used similar methods to investigate the composition of marine bacterioplankton communities. Post-sorting genomic techniques were used in both cases for additional sample characterization (62,63).…”

Section: Combining Fish and Flow Cytometrymentioning

confidence: 99%

“…Kalyuzhnaya et al (62) used this approach to examine methanotrophic bacteria from lake sediments and Sekar et al (63) used similar methods to investigate the composition of marine bacterioplankton communities. Post-sorting genomic techniques were used in both cases for additional sample characterization (62,63). Food applications of FISH-FC include the use of a Pseudomonas-specific DNA probe to detect this spoilage organism in milk (64) and detection of Salmonella spp.…”

Section: Combining Fish and Flow Cytometrymentioning

confidence: 99%

Methods for Whole Cell Detection of Microorganisms

Brehm-Stecher

2008

ACS Symposium Series

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Microbes are ubiquitous, and can be found occupying nearly every imaginable niche on Earth. These include organic and inorganic surfaces, interfacial boundaries and within macroscopically solid matrices, such as the pore space of rocks. Because phylogenetically divergent microbes may be visually indistinguishable, understanding the species distribution and ecological significance of environmental microbes requires diagnostic tools that extend beyond simple phenotypic description. Methods for microbial diagnostics can be divided into two broad categories: cellular and acellular. Acellular techniques, such as the polymerase chain reaction or certain immunoassay formats may be effective at detecting molecular, structural or biochemical targets associated with specific cell types, but this information is provided out of its "natural", and arguably most meaningful context -that of the individual microbial cell. In contrast, cellular methods have the potential to preserve an abundance of valuable information. Apart from molecular identity, this includes information regarding cell morphology and other physiological characteristics, cell number and distribution within a sample, and physical or spatial associations with other cell types. The aim of this chapter is to provide an overview of whole cell methods for microbial detection, including both existing approaches and those still in development. The tools described here are expected to find wide application for the detection of microbes on surfaces or within complex matrices across a number of parallel or allied fields, including environmental, food and clinical microbiologies.

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Flow Sorting of Marine Bacterioplankton after Fluorescence In Situ Hybridization

Cited by 139 publications

References 61 publications

Prokaryotic community structure and heterotrophic production in a river-influenced coastal arctic ecosystem

Prokaryotic community structure and heterotrophic production in a river-influenced coastal arctic ecosystem

Review: Microbial biocenoses in pristine aquifers and an assessment of investigative methods

Methods for Whole Cell Detection of Microorganisms

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